A phylogenetically distinctive and extremely heat stable light-driven proton pump from the eubacterium Rubrobacter xylanophilus DSM 9941T

Kanae Kanehara,Susumu Yoshizawa,Takashi Tsukamoto,Yuki Sudo

doi:10.1038/srep44427

Kanae Kanehara, Susumu Yoshizawa + Show 2 more

Open Access

https://doi.org/10.1038/srep44427

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Journal: Scientific Reports	Publication Date: Mar 14, 2017
Citations: 26	License type: open-access

Affiliation: Okayama University, The University of Tokyo

Abstract

Rhodopsins are proteins that contain seven transmembrane domains with a chromophore retinal and that function as photoreceptors for light-energy conversion and light-signal transduction in a wide variety of organisms. Here we characterized a phylogenetically distinctive new rhodopsin from the thermophilic eubacterium Rubrobacter xylanophilus DSM 9941T that was isolated from thermally polluted water. Although R. xylanophilus rhodopsin (RxR) is from Actinobacteria, it is located between eukaryotic and archaeal rhodopsins in the phylogenetic tree. Escherichia coli cells expressing RxR showed a light-induced decrease in environmental pH and inhibition by a protonophore, indicating that it works as a light-driven outward proton pump. We characterized purified RxR spectroscopically, and showed that it has an absorption maximum at 541 nm and binds nearly 100% all-trans retinal. The pKa values for the protonated retinal Schiff base and its counterion were estimated to be 10.7 and 1.3, respectively. Time-resolved flash-photolysis experiments revealed the formation of a red-shifted intermediate. Of note, RxR showed an extremely high thermal stability in comparison with other proton pumping rhodopsins such as thermophilic rhodopsin TR (by 16-times) and bacteriorhodopsin from Halobacterium salinarum (HsBR, by 4-times).

Highlights

Rhodopsins are proteins that contain seven transmembrane domains with a chromophore retinal and that function as photoreceptors for light-energy conversion and light-signal transduction in a wide variety of organisms
A putative rhodopsin protein R. xylanophilus rhodopsin (RxR) was discovered in the thermophilic eubacterium R. xylanophilus in 2014 (GenBank ID: ABG04982)
RxR is produced in a eubacterium, its amino acid sequence is most closely related to the archaeal proton pumps (44% identity, 71% similarity in comparison with H. salinarum bacteriorhodopsin (HsBR) and 49% identity, 81% similarity in comparison with AR3) and eukaryotic proton pumps (37% identity, 68% similarity in comparison with Leptosphaeria rhodopsin (LR)), but not to eubacterial proton pumps (21% identity, 59% similarity in comparison with PR, 23% identity, 63% similarity in comparison with XR, and 25% identity, 59% similarity in comparison with thermophilic rhodopsin (TR))

Summary

Introduction

Rhodopsins are proteins that contain seven transmembrane domains with a chromophore retinal and that function as photoreceptors for light-energy conversion and light-signal transduction in a wide variety of organisms. We characterized the purified RxR spectroscopically and its properties were compared with those of other microbial proton pumping rhodopsins.

Results

Conclusion